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Exome sequencing for diagnostics         Christian Gilissen   Radboud University Nijmegen Medical Center        Department...
Published exome sequencing studies                                Gilissen et al. Genome Biology, 2011
Strategies to prioritize variants from exome studies                                      SETBP1 – Schinzel Giedion syndro...
DiagnosticsCan we use sequencing technologies for the  diagnosis of genetically heterogeneous  diseases were current tests...
Targeted versus whole exome• Targeted:   • Low chance of incidental findings   • Easy analysis   • Cheap (?)• Exome:   • U...
ApproachesGene package approach   • Exome sequence a single individual with a specific     indication       •In vitro enri...
How to implement Exome sequencing for diagnostics 1.Increase diagnostic yield 3.Prevent incidental findings 5.Quality cont...
Results• Proof of Concept: 50 samples for 5 genetically  heterogeneous disorders
Analysis1.Quality control3.Coverage of gene packages5.Interpretation7.Visualization
1. Quality Control• QC statistics• Gender match• 12 SNP pyrosequencing test• Trio concordance
2. Coverage of disease gene package
2. Disease gene coverage
Disease gene coverage - Agilent SureSelect version 4
% reads showing          Known                 variant               SNP                                              HGMD...
3. Interpretation•   Automated annotation per indication    •  Database of known disease genes•   Automated prioritization...
4. Visualization
Results – variants requiring attention
Results – diagnostic yield?• Exact diagnostic yield is still being calculated• Preliminary results indicate that:   • Yiel...
Conclusion• We designed and implemented an analysis for exome  sequencing based genetic diagnosis of genetically  heteroge...
Acknowledgments• Rick de Reuver• Marisol del Rosario• Nienke Wieskamp• Yannick Smits• Joris Veltman• Marcel Nelen• Hans Sc...
Prioritization of variants causing genetic disease• Number of coding variants:   ~ 12,000• Private* non-synonymous   varia...
Exome sequencing for disease gene identification and patient diagnostics, Genomic Disorders Nijmegen, Radboud University N...
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Exome sequencing for disease gene identification and patient diagnostics, Genomic Disorders Nijmegen, Radboud University Nijmegen Medical Centre, Christian Gillisen, Copenhagenomics 2012

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Exome sequencing for disease gene identification and patient diagnostics

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Exome sequencing for disease gene identification and patient diagnostics, Genomic Disorders Nijmegen, Radboud University Nijmegen Medical Centre, Christian Gillisen, Copenhagenomics 2012

  1. 1. Exome sequencing for diagnostics Christian Gilissen Radboud University Nijmegen Medical Center Department of Human Genetics c.gilissen@gen.umcn.nl
  2. 2. Published exome sequencing studies Gilissen et al. Genome Biology, 2011
  3. 3. Strategies to prioritize variants from exome studies SETBP1 – Schinzel Giedion syndrome Hoischen et al. Nat gen. 2010 WDR35 – Sensenbrenner syndrome Gilissen et al. AJHG 2010 DYNC1H1 etc. – Intellectual disability Vissers et al. Nat gen. 2010 SERPIN1F – Osteogenisis Imperfecta Becker et al. AJHG 2011 ASXL1 – Bohring-Opitz syndrome Hoischen et al. Nat gen. 2011 IMPAD1 - Chondrodysplasia Vissers et al. AJHG 2011 ACTB/ACTG – Baraitser-Winter syn. Riviere et al Nat gen. 2012 ABCC9 - Cantu syndrome Van Bon et al. AJHG 2012Gilissen et al. EJHG, 2012
  4. 4. DiagnosticsCan we use sequencing technologies for the diagnosis of genetically heterogeneous diseases were current tests have a low diagnostic yield?
  5. 5. Targeted versus whole exome• Targeted: • Low chance of incidental findings • Easy analysis • Cheap (?)• Exome: • Unbiased, not dependent on a gene set • Uniform approach for all diseases • Synergy
  6. 6. ApproachesGene package approach • Exome sequence a single individual with a specific indication •In vitro enrichment for known disease genes •Prioritization for obvious candidates outside of known genesDe novo approach • Exome sequence patient-parent trio •In vitro enrichment for known disease genes •De novo candidates •X-linked and recessive candidates •Prioritization for obvious candidates outside of known genes
  7. 7. How to implement Exome sequencing for diagnostics 1.Increase diagnostic yield 3.Prevent incidental findings 5.Quality control of sample 7.Easy and standardized interpretation
  8. 8. Results• Proof of Concept: 50 samples for 5 genetically heterogeneous disorders
  9. 9. Analysis1.Quality control3.Coverage of gene packages5.Interpretation7.Visualization
  10. 10. 1. Quality Control• QC statistics• Gender match• 12 SNP pyrosequencing test• Trio concordance
  11. 11. 2. Coverage of disease gene package
  12. 12. 2. Disease gene coverage
  13. 13. Disease gene coverage - Agilent SureSelect version 4
  14. 14. % reads showing Known variant SNP HGMD Kegg GO Exonic / Splice site OMIM Protein Mouse domainsphenotypes Other exomes Grantham Conservation
  15. 15. 3. Interpretation• Automated annotation per indication • Database of known disease genes• Automated prioritization• Graphical user interface • Stand-alone web-application • Flexible with respect to input files • Prevent unrelated findings • Flexible with respect prioritization • Allow for quality control
  16. 16. 4. Visualization
  17. 17. Results – variants requiring attention
  18. 18. Results – diagnostic yield?• Exact diagnostic yield is still being calculated• Preliminary results indicate that: • Yield varies considerably per gene package • Considerably higher than with current diagnostic tests • E.g. blindness package: > 40% positive reports
  19. 19. Conclusion• We designed and implemented an analysis for exome sequencing based genetic diagnosis of genetically heterogeneous disorders. • Minimizes risk of incidental findings • Allows for strict quality control • Standardized interpretation
  20. 20. Acknowledgments• Rick de Reuver• Marisol del Rosario• Nienke Wieskamp• Yannick Smits• Joris Veltman• Marcel Nelen• Hans Scheffer• Genomic disorders group• DNA Diagnostics Nijmegen
  21. 21. Prioritization of variants causing genetic disease• Number of coding variants: ~ 12,000• Private* non-synonymous variants: ~ 150-200*Not detected in dbSNP or other controldata. How to identify the 1 causative variant?

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